JPS627966B2 - - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to an apparatus for measuring the concentration of particles suspended in a fluid, and more particularly to a device for measuring the concentration of particles suspended in a fluid, and in particular for measuring the concentration of particles of at least two types, one of which has a readily detectable or standard concentration. The present invention relates to an apparatus for measuring the concentration of particles of a first type in a fluid in which particles of a first type are suspended. The invention is particularly suitable for determining the number corresponding to the number of particles per unit volume of whole blood. Such a number is often referred to as platelet density or concentration. The concentration may be expressed in terms of particles per unit volume or in terms of hematocrit or fractional volume.

Blood testing is one of the most common and most frequently used methods of medical diagnosis. Normal standard counts of red blood cells, white blood cells, platelets, and other constituent particles of blood, as well as the statistical distribution of such counts among healthy individuals, have been established. Such counts are usually expressed as counts per unit volume, or concentration. Deviations from these standard values often indicate the presence of an infectious disease or other medical problem.

The number of platelets per unit volume is an important means for clinical diagnosis in blood tests. Substantial deviations in the number of platelets per unit volume of blood from normal values often indicate the presence of disease. For example, tuberculosis often produces an increased number of platelets, and other acute infectious diseases may produce less than normal numbers. A marked decrease in platelet count is often indicative of acute leukemia. A moderate decrease in platelet count indicates an adverse effect of the drug.

Platelets are small and more numerous than red blood cells. Typically, normal human blood contains only about 5 million red blood cells per microliter of whole blood and about 250,000 platelets per microliter. Red blood cells are usually about 8 in diameter
They are micron biconcave discs, and platelets are round, oval, or rod-shaped particles about 2.5 microns in diameter.

Platelets are difficult to count in the presence of red blood cells because of their small size and low number compared to red blood cells. However, special processes that separate platelets or destroy red blood cells before performing a platelet count not only add delay and expense to the counting process, but also reduce the reliability of the process. , it is desirable to count platelets in the presence of red blood cells.

In previous indirect methods of detecting platelet concentration, platelets and red blood cells are deposited on a microscope slide until a certain minimum number of red blood cells are counted in a selected area. counted at the same time. The number of platelets is then obtained by the ratio of platelets to red blood cells and the more easily determined concentration of red blood cells, which concentration is determined from counts obtained from a known volume of the same sample of blood. It is.

In one such process, one part of whole blood is typically diluted to ninety-nine parts of a physiological saline solution, such as ISOTON, a product of Coulter Electronics Company. The mixture is then transferred to a glass slide and after 15 to 45 minutes the slide is examined under a microscope. Sometimes counting is carried out with the aid of a counting chamber containing several small square blocks. The number of particles counted in, say, 10 such blocks is multiplied by a constant based on the dilution ratio and block size. Platelets are visually counted simultaneously with red blood cells until 1000 red blood cells are counted in a common area of the glass slide. The number of platelets counted is, for example, 68. The number of red blood cells per unit volume, that is, the concentration of red blood cells, is already known.
Alternatively, it can be determined by other commonly required tests of blood. For the purpose of explanation,
Concentration of red blood cells per microliter
Let's assume that there are 4480000 pieces. This represents the actual concentration measured in the sample to which the invention was applied.

Platelet concentration P _D is the ratio of counted platelets to counted red blood cells, RBC _D
It is equal to the profit.

In this example, P _D = RBC _D x 68/1000 = 4480000 x 0.068 = 304640 platelets/microliter. This has the advantage that the counting process can be carried out irrespective of the volume of the sample, as long as it is large enough to give a fairly accurate platelet concentration indication. This and other methods of counting platelets were published by WB Saunders Company in 1969.
Publisher, Israel Davidsohn and John Bernard
Todd−Sanford Clinical Diagnosis by Henry
by Laboratory Methods, 14th edition, pages 157-161.

These traditional indirect methods of counting platelets and red blood cells simultaneously are either very time consuming or relatively inaccurate due to the small number of particles counted. For example, in the above explanation,
Even a single platelet error in platelet counting can cause a 1.5% error in the platelet concentration count.
Furthermore, previous platelet concentration calculation methods based on simultaneous counting of red blood cells and platelets require algebraic processing following completion of the counting process.

Electronic particle counters allow particles in blood to be counted in a very short time and with greater accuracy than was previously possible using microscopic techniques.

The general concept of electronically counting particles of various types suspended in a liquid sample and having distinguishable physical characteristics in known quantities through a sensing transducer is well known. be. See, for example, Coulter patent no. 2,656,508.

However, to the applicant's knowledge, electronic particle counting has not been used to date for indirect methods of simultaneously counting red blood cells and platelets to determine the platelet count concentration of a variable volume of diluted whole blood. Not applied. Furthermore, to the best of Applicant's knowledge, prior to the invention described herein, the algebraic processing required to calculate platelet counts per unit volume following coincidence counting of red blood cells and platelets of a blood sample has not been disclosed. There is no known way to avoid this.

Particle concentration or density in a liquid medium is usually expressed in terms of counts per unit volume, but may also be expressed in other ways, such as percent volume. In particular, in the analysis of blood, routine measurements of both the red blood cell count and the hematocrit, ie, the percentage of blood volume occupied by red blood cells, of a sample are often performed.

Accurate reference values for hematocrit are usually produced by known centrifugation processes.

In known electronic processes for measuring hematocrit, the pulses to be counted are first weighted in proportion to the volume of each particle being counted. The weighted pulses are then summed or integrated to produce a signal proportional to the percent volume of liquid occupied by particles.

Although the invention can be applied in other ways, for the sake of simplicity it will be specifically described in terms of density or concentration expressed in counts per unit volume. It is therefore to be understood that the broad concepts underlying the present invention can also be applied to density or concentration measured and expressed in other ways than counts per unit volume.

The primary object of the present invention is to provide an improved device for measuring the platelet concentration of a variable volume blood sample by simultaneously electronically measuring the amount of red blood cells and platelets in the blood sample. .

It is also an object of the present invention to provide a device for determining the number of platelets per unit volume of a blood sample of variable volume without requiring algebraic processing of red blood cell and platelet counts.

Yet another object of the invention is to simultaneously electronically count platelets and red blood cells in a diluted variable volume of blood, one corresponding to the red blood cell count and the other.
It would be better to provide a device that generates a separate count corresponding to the platelet count. Means for automatically terminating the counting process when the red blood cell count reaches a predetermined value so that the displayed platelet count directly indicates the number of platelets per unit volume, eliminating the need for algebraic processing of the data. It is also an object of the present invention to provide such a device.

Yet another object of the present invention is to provide a circuit that reduces the possibility of accidental pulse generation in response to spurious signals such as noise, and that the smaller of two particles detected at about the same time is detected first. The purpose is to improve the accuracy of particle counting by using a circuit that counts the first particle even if the particle is detected at the beginning of the particle detection process.

Yet another object of the invention is to automatically terminate the counting process when certain conditions occur.

Yet another object of the present invention is to provide an apparatus that automatically takes into account error due to coincidence effects in the number of pulses generated by passing a particle stream through a particle sensing transducer. It is.

In particular embodiments of the invention described in detail herein, circuitry is provided to discriminate the electrical signals generated by the detection of platelets from the electrical signals generated by the detection of red blood cells, and the number of red blood cell pulses counted is providing a circuit for separately counting electrical pulses generally corresponding to detected platelets and electrical pulses generally corresponding to detected red blood cells until a value is reached and automatically stopping the platelet counting process; Accordingly, a pulse count that generally corresponds to the detected platelets or a platelet count that corresponds per unit volume can be automatically displayed.

A very advantageous feature of the preferred embodiment of the present invention is the application of modern high speed electronic counting devices that directly measure the number of platelets per unit volume, which would otherwise normally be required. The object of the present invention is to provide circuit means that can eliminate algebraic processing.

Another highly advantageous feature of the preferred embodiment of the invention is the provision of a count termination means that can be set at a known red blood cell count concentration which automatically stops the electronic count.

Yet another very advantageous feature of the present invention is that the measurement is independent of sample volume so that the accuracy of sample dilution does not have to be exact. This feature is particularly advantageous as some embodiments of the invention prefer to use a sheath to focus or confine the sample flow to the center of the particle transducer, as discussed below.

Other objects, features and advantages of the invention will become apparent from the following detailed description taken in conjunction with the accompanying drawings.

GENERAL DESCRIPTION Referring now to FIG. 1, a platelet counting device collects a diluted sample of blood to generate a flow of electrical signals with amplitudes correlated to certain physical properties of the detected particles. is generally shown comprising a particle sensing transducer 10 through which a particle sensing transducer 10 is flown.

The difference in size between red blood cells and platelets provides relatively convenient physical characteristics for distinguishing these particles. There are a number of known methods and devices for generating electrical signals whose amplitude depends on the size of particles suspended in a fluid, such as red blood cells or platelets suspended in diluted whole blood. One such known method is to change the relative electrical impedance between a pair of electrodes suspended in a conductive fluid as the particle passes through a hole slightly larger than its diameter. Other methods are also known, including the use of photodetectors.

Particle sensing transducer 10 may be any such known device as long as it generates an electrical signal whose characteristics are based on the size of the particle being detected. In the embodiments of the invention described below, the particle sensing transducer is of a type that generates an electrical signal having an amplitude substantially proportional to the size of the particle being detected.

Additionally, the particle sensing transducer should preferably avoid problems associated with particles recirculating after passing through the sensing region of the transducer. If these types of problems are not resolved, they will make it difficult or even impossible to accurately count particles of different sizes suspended in the same sample. For example, one such preferred transducer is
Hogg patent no. 3902115. Another suitable transducer was installed in March 1976.
Scientific American, Volume 234, Issue 3, pages 108-117, “Fluorescence-Activated
It is described in a paper titled "Cell Sorting".
Yet another transducer that solves the above problems and is suitable for the present invention was published in July 1973 in the Medical and
Schultz in Biological Engineering, pp. 447-454.
and Thom, “Electrical Sizing and Counting
of Platelets in Whole Blood”.

In practice, the number of pulses generated by a transducer of the above type when a stream of particles passes through the sensing region is less than the number of particles flowing into the sensing region. One reason for this is that two or more particles may flow into the sensing region in close proximity to each other, producing superimposed pulses such as two partially superimposed pulses, or producing only a single pulse. This is for the purpose of Therefore, when two types of particles of different size are to be detected, the presence of one particle is likely to generate a pulse that masks the pulse due to the presence of the other particle. The present invention reduces errors that may be caused by such obscuring and other matching effects. However, in order to simplify the explanation of the invention, it will first be assumed that the particles flow into the sensing region without such interference, and then that many such obscuring effects occur but are eliminated or eliminated. Embodiments of the present invention are described in which the errors likely to occur due to coincidence are at least reduced, and it is shown that all embodiments of the present invention generally significantly reduce, although not completely eliminate, the errors that are likely to occur due to matching effects. In either case, the best practice of the invention is such that errors in the counting pulses that occur in coincidence events are virtually non-trivial.

It is well known that red blood cells and platelets are not uniformly sized. In reality, the sizes of each type of particle are widely distributed, but they overlap only to a small extent. For convenience of explanation, unless otherwise indicated, the present invention will be described assuming that this overlap is negligible.

The signal generated by the particle sensing transducer 10 is typically amplified before it can be used in readily available commercially available components such as those operating in the 0 to 15 volt voltage range. I need. A simple AC amplifier 12 with a voltage gain of about 2000 and a bandwidth of about 50 KHz is typically used for variable impedance particle sensing transducers. The direct current (DC) voltage generated by the particle-sensing transducer provides no information regarding particle size and may interfere with the operation of the discriminator circuitry used to distinguish between particles of different sizes, as described below. Since this is a win-win situation, it is preferable to use an AC amplifier.

High threshold detector 16 and low threshold detector 18 distinguish pulses corresponding to platelets from pulses corresponding to red blood cells and distinguish such pulses from low level noise that may be counted accidentally. used for

To simplify the threshold detection operation, the lower limits of the platelet and red blood cell electrical signals are clamped to ground. This clamp is the baseline return circuit 14.
achieved by. The peak detector 20 is timed to detect peaks in the platelet and red blood cell electrical signals and to sample the output signals of the high and low threshold detectors approximately at the time the peak amplitude of the electrical signal occurs. Serves as a means for giving signals.

The combination of AC amplifier 12 and baseline return circuit 14 also serves to make the counting operation relatively insensitive to baseline instability in the particle sensing transducer. Baseline instability, ie, random changes in the fixed operating voltage, can occur in fluid conductivity measurement devices, for example due to the formation of air bubbles.

Pulse classification unit 22 receives the output signals of high threshold detector 16, low threshold detector 18, and peak detector 20, respectively, and, in response, determines the type of particle detected, i.e., red blood cell or A pulse is generated on one of the two lines 33 and 35 corresponding to the platelets.
Therefore, the pulse classification unit 22 converts each electrical signal generated by the detection of particles into a pulse corresponding to red blood cells or a pulse corresponding to platelets;
These pulses are then transmitted on a separate line, namely a first pulse line 33 dedicated to red blood cell pulses.
and a second pulse line 35 used exclusively for platelet pulses to the counter.

As shown on the right side of FIG. 1, two pulse lines 33 and 35 from pulse classification unit 22
are connected to separate counters. Platelet pulse line 35 is connected to platelet pulse counter 24 and red blood cell pulse line 33 is connected to red blood cell pulse counter 28. Since each counter is activated only by pulses corresponding to a particular type of particle designated, the count produced by each counter corresponds only to the number of particles of that type. Thus, the red blood cell pulse counter counts the number of red blood cells that have passed the particle sensing transducer 10, and the platelet pulse counter 24 counts the number of platelets that have passed the particle sensing transducer 10.

As shown on the right side of FIG. 1, the platelet pulse counter 24 is connected to a platelet display unit 26 where the platelet counts generated are displayed and thus viewed by the user. On the other hand, the output of the red blood cell pulse counter 28 is connected to a register in the form of a thumb-operated wheel switch array 30, which array 30 is connected to the already known or assumed number of red blood cells per unit volume of the blood sample being tested. is set to a predetermined count that typically corresponds to . When the output of the red blood cell pulse counter 28 corresponds to the preset count set in the thumb-operated wheel switch array 30, a count end signal is generated.
A STOP is generated, which automatically and immediately terminates the counting process occurring in the red blood cell pulse counter 28 and platelet pulse counter 24. The number of red blood cells counted is equal to the preset value set in the thumb wheel switch arrangement 30, and the platelet pulse counter simultaneously displays the number of platelets counted. The platelet display then corresponds to the number of platelets per unit volume in the sample used to count red blood cells.

Although the embodiment of FIG. 1 is suitable for practicing the invention, it may be considered desirable under certain laboratory conditions and the preferred embodiment of the circuit of the invention shown in FIGS. 2A and 2B. There are certain additional benefits as provided by the embodiments. For example, the circuit of FIG. 2A uses a baseline return circuit that also acts as a high pass filter to reduce the sensitivity of the circuit to low frequency noise signals such as 60 Hz AC. Furthermore, for example, the second
Embodiments of the invention using the circuit of Figure A are highly unlikely to generate an erroneous number of pulses in response to a noise signal superimposed on the sense transducer output signal. Furthermore, the circuit of FIG. 2A is better suited for generating platelet pulses corresponding to detected platelets just before and nearly coincident with red blood cells. The circuit of FIG. 2A thus avoids the obscuring effects inherent in large red blood cells during the counting process. These and other advantages of the circuits of FIGS. 2A and 2B will become more fully apparent from the detailed description of various embodiments of the invention.

In the best mode of practicing the invention, the end-of-count signal STOP is used to terminate the two counting operations being performed. However, as long as the platelet count generated when the preset red blood cell count is reached is made available to the user, the present invention will work even if only the platelet counting operation is terminated and even if the counting operation is not stopped. The main effect will be achieved.

In a preferred embodiment of the device of the invention, the red blood cell concentration of the sample under test is known and the sensing transducer employs a sheath focusing device. In a flow system most suitable for use with the present invention, i.e., one that involves accurate measurements of particle size and counts, a focusing device such that a core of sample fluid flows into the center of a sheath of particle-free fluid. It is desirable to use Such a device can project the sample precisely along the center of the sensing area. However, in such devices, it is very difficult to size the relative volumes of the sample and sheath because the sheath focusing device causes uncontrolled sample dilution. This uncontrolled dilution eliminates sample-flow volume measurements. This is because the measured flow volume is for an unknown size of sample and sheath, and the volume and flow rate are typically about 50 times larger in such a focusing device than in an unfocused device. This is because it is also small.

Sample volumes of 10 to 30 microliters and flow rates of 0.1 to 0.5 microliters per second are typical for focusing devices.
Devices for measuring such small volumes and flow rates are difficult to make accurate, reliable, and inexpensive. However, using the present invention and knowing in advance the particle density or concentration of one of the constituent particles of a sample, the particle density or concentration of another type of constituent particle of the sample can be determined. The only condition is that the different types of particles can be distinguished from each other by certain physical properties. Furthermore, the concentration of particles can be detected even though the dilution ratio is not controlled and the sample flow rate is unknown and small.

In the present invention, prior knowledge of one of the constituent particles of the sample under test is typically used. If such prior knowledge is not available,
The same transducer can initially be used without a sheath to accurately measure the density or concentration of one constituent particle before proceeding with the simultaneous counting of all constituent particles of the sample.

Additionally, if no prior knowledge of the constituent particles of one of the samples is available, the present invention provides for adding a second sample containing a known concentration of physically distinguishable particles to the sample under test. A further step of mixing is intended. Although the initial sample mixture must be accurate, subsequent sampling and dilution of this mixture may not be accurate. Therefore, the above-described problem of simultaneously counting particles of different sizes while using a front sheath is overcome in this embodiment of the invention. Second
The sample particles may be any particles suitable for sensing in the sensing region of the transducer, so long as they can be distinguished from particles of unknown concentration in the sample. Synthetic particles of known size and known concentration are typically suitable for such purposes. For example, polyethylene or polystyrene beads manufactured by Dow Corning, Midland, Michigan, USA, are suitable synthetic particles for blood sample measurements performed in accordance with the present invention.

DETAILED DESCRIPTION In this detailed description of the invention, a method for distinguishing between pulses generated as a result of detecting platelets and pulses generated as a result of detecting red blood cells and counting such pulses is described. The apparatus and method are described in sufficient detail to enable those of ordinary skill in the art to make the apparatus and practice the method in the best mode currently contemplated. The particle sensing transducer 10 and AC amplifier 12 shown in FIG. 1 are each well known, and neither requires further explanation. Note only that the flow of a sample volume of whole blood through particle sensing transducer 10 produces an electrical signal at the output of AC amplifier 12, such as that shown by waveform WA' in FIG. This waveform WA' corresponds to the electrical signal appearing at circuit point A', ie, the output signal of the AC amplifier 12 when red blood cells and then platelets are detected by the particle sensing transducer 10.

In FIG. 1, the shape of waveform WA' corresponds to the change in resistance that occurs between the electrodes of transducer 10 as sample flows through transducer 10. As shown, pulses of different heights are generated depending on whether red blood cells or platelets are detected. In one variation of the invention, this electrical signal is differentiated and the relative heights of the pulses in the resulting waveform are again detected. In the illustrated waveform WA', large pulses correspond to red blood cells and small pulses correspond to platelets. After differentiation, the relative heights of the pulses in this differentiated signal are preserved, such that small pulses correspond to platelets and large pulses correspond to red blood cells. Therefore, different pulse characteristics corresponding to different particles may be used in the practice of the present invention. However, for the sake of brevity, the invention will be described in detail with reference only to electrical signals of the type represented by the particular waveform WA'.

As shown in FIG. 1, the baseline return circuit 14 includes a series capacitor 11, a diode 13, and a resistor 15 connected in parallel between point A and ground G. As mentioned above, the purpose of this baseline return circuit 14 is to
The purpose is to restore the 0 Volt DC baseline to the AC coupled output signal of the AC amplifier 12 so that it can be used more conveniently.

The output signal of the baseline return circuit 14 for the input signal corresponding to the waveform WA' corresponds to the waveform WA, or two pulses, the larger of which corresponds to the earlier detected red blood cells and the smaller one to the later detected red blood cells. Corresponding to platelets, the common baseline is approximately 0 volts DC. The reference time “to” of each waveform is the same, and all waveforms WA′, WA, WB,
It is intended to give a rough indication of the relative temporal relationships of WC, WD, WE and WF.

A high threshold detector 16 and a low threshold detector 18 are each connected to a voltage comparator 1.
7 and 19 are used. These two threshold detectors 16 and 18 are identical in all respects except for the voltages applied to potentiometers 21 and 23, respectively. The potentiometer 21 of the high threshold detector 16 is connected at its first end to a power supply of approximately 3 volts DC and at its second end to a power supply of approximately 5 volts DC, thus The voltage at the arm of meter 21 relative to ground can be varied between 3 and 5 volts DC. On the other hand, the potentiometer 2 of the low threshold detector 18
3 is connected at its first end to a power supply of approximately 1 volt DC and at its second end to ground, thus making the voltage at the arm of potentiometer 23 relative to ground 0 volts DC and 1 volt DC. can be changed between. In this case, the two voltage ranges do not overlap.

Each threshold detector 16 or 18
uses a voltage comparator which is a high gain amplifier 17 or 19 respectively, the output signal of which is, as the case may be,
The higher limit is pushed when the input voltage applied to the positive input line of terminal 2 exceeds the reference voltage applied to the negative input line of terminal 3, and the lower limit is pushed when a reaction occurs. One such voltage comparator is the “Linear
National Integrated Circuit”
Semiconductor publication (1975 edition) page 3
-National listed in 21 to 3-25
Semiconductor Model LM311 voltage comparator/buffer.

Terminals 4 and 8 of each voltage comparator are connected to -15 volts DC and +15 volts DC, respectively, and terminal 1 is connected to ground G. The output terminal 7 of each comparator is connected to the input terminal 2 by a feedback resistor R _f .
This feedback configuration including input resistance R _i is:
The voltage difference between the positive and negative input lines increases the precision with which the output signal is pushed from one limit to the other.

In FIG. 1, waveform WB represents the electrical signal generated at point B in response to application of the waveform WA input signal corresponding to circuit point A to detector 16. In FIG. This waveform WB is a signal pulse of a width corresponding to the portion of the red blood cell detection signal that exceeds the high threshold level set in high threshold detector 16 by potentiometer 21. The electrical signal corresponding to the detection of platelets does not produce a corresponding output signal at point B. This is because the threshold detector 16 has a high platelet pulse amplitude.
This is because it is smaller than the red blood cell discrimination level.

Also, in FIG. 1, waveform WC represents the output signal of low threshold detector 18.
Waveform WC contains two electrical signals or pulses, one generated by the red blood cell signal above the platelet discrimination level and the other generated by the platelet signal above that level. It is. Since both electrical signals generated by the detection of each particle exceed the platelet discrimination level,
The output of the low threshold detector 18 has 2
Two pulses are generated. Note that the pulse corresponding to the red blood cell electrical signal is large in width. This large width occurs because the time the red blood cell electrical signal exceeds the platelet discrimination level is greater than the time the platelet signal exceeds this level.

Peak detector 20 includes a voltage comparator 25, which is a National Semiconductor LM311, the same voltage comparator suitable for use in threshold detectors 16 and 18. Terminal 3 of this voltage comparator 25, ie the negative voltage input line, is connected to ground reference G. The input line to the positive voltage terminal of voltage comparator 25 is connected to series capacitor 27.
and a parallel resistor 29 connected to ground G. This capacitor and resistor form a simple differentiator circuit which, as is well known, produces a non-zero output signal only when the input signal changes, and a zero output signal when the input signal is constant. It just happens. Since the input signals for the detected red blood cells and platelets have a positive slope, a zero slope, and a negative slope in this order, the differentiating circuit receives a positive signal during the transient period of the positive slope of the input signal. It produces a zero output during zero slope times and a negative signal during negative slope transient times. The output signal of voltage comparator 25 is shown in FIG. 1 as waveform WD, which corresponds to the signal occurring at point D with respect to input waveform WA'. As shown in waveform WD, the output signal of peak detector 20 is a positive signal during the positive slope time of the red blood cell pulse, and a positive signal during the positive slope time of the platelet pulse, and all other times. It is zero during the time. The purpose of the peak detector in this embodiment is to provide a timing signal that indicates approximately when the peak of the electrical signal generated by particle detection occurs. National
Using offset balancing of the type shown on pages 3-21 of the aforementioned publication of Semiconductor Inc.,
The voltage comparator can be adjusted so that it produces an output signal voltage when the differential input voltage is slightly above zero voltage.

High threshold detector 16, low threshold detector 18, and peak detector 2
The purpose of 0 is to generate a signal that is used to distinguish between the electrical signals generated by the detection of red blood cells and the electrical signals generated by the detection of platelets, and to send the separately counted pulses to separate lines 33 and 33. 35, one set of pulses on the first line 35 corresponds only to platelets and another set of pulses on the second line 33 corresponds only to red blood cells. The conversion of the signals generated at detectors 16, 18 and 20 into separate pulses applied to separate lines 33 and 35 is accomplished in pulse classification unit 22.

The pulse classification unit 22 is an AND gate AG.
1, two logic inverters IN1 and IN2, and a D-type flip-flop FF1. An AND gate produces a 1 or true output signal (eg, +5 volts DC) only when a 1 or true input signal is simultaneously applied to each of its input lines. The AND gate produces a zero or false output signal (eg, 0 volts DC) at other times. A logic inverter produces a one or true output signal when its input signal is zero or false, and produces a zero or false output signal when the input signal is one or true. The Q terminal of a D-type flip-flop produces a zero or false output signal as long as a zero or false signal is applied to its reset terminal R. However, when the signal applied to the reset terminal R is a 1 or true signal, the Q terminal will cause the data terminal D to appear on each positive edge of the clock signal CL.
generates a signal that has the same logic state as the logic state of the signal applied to it.

The output signals of the high threshold detector 16 and the peak detector 20 are each connected to an AND gate AG1.
is applied to the two input lines of Thus, AND gate AG1 produces one output signal only when the signals appearing at points B and D are simultaneously positive voltages. Therefore, when the peak of the detected signal approximately occurs, when the amplitude of this detected signal exceeds the high threshold level set in the high threshold detector 16, corresponds to the detection of red blood cells. The pulse is the red blood cell pulse line 3.
3 is generated at the output of the AND gate AG1.

As shown in FIG.
2 outputs the inverted output signal WB of the high threshold detector 16 to a flip-flop.
Inverter IN1 is used to apply data to the data input D of FF1. Peak detector output signal
Flip the inverted version of WD - Flop FF
Another inverter IN2 is used to supply the clock terminal CL of clock 1. Also, as shown, a low threshold detector 18
The output signal WC of is applied to the reset terminal R of flip-flop FF1. The output terminal Q of flip-flop FF1 is directly connected to the platelet pulse line 35, so that the Q output of flip-flop FF1 determines the state of the signal appearing on the platelet pulse line 35.

If the detected input signal WA does not exceed the low threshold level, the reset terminal R of flip-flop FF1 is held false, preventing the output signal WF of terminal Q of flip-flop FF1 from becoming true. . On the other hand, if the signal at the output of low threshold detector 18 becomes true due to the detected input signal exceeding the low threshold level, reset terminal R of flip-flop FF1
is set to the true level. Then, while a 1 or true signal is applied to the reset terminal R of flip-flop FF1, the output terminal Q
occurs when a positive going signal is applied to the clock terminal CL of flip-flop FF1.
Generates an output signal having the same logic state as the signal applied to data terminal D.

If the detected signal WA exceeds the low threshold level of low threshold detector 18 and also exceeds the high threshold level of high threshold detector 16, flip-flop FF1 is enabled. , and the input signal applied to the data terminal D of flip-flop FF1 becomes false. Therefore, when peak detector 20 produces an output signal at or about the time this detected signal peak occurs, flip-flop FF1 is clocked and the data input of flip-flop FF1 is clocked. An output signal appears at output terminal Q with a false logic state corresponding to the false logic state applied to terminal D. On the other hand, as described above, AND gate AG1 generates signal WE having a true logic state. Therefore, when red blood cells are detected, a pulse corresponding to the presence of detected red blood cells is applied to the red blood cell line 33 and no pulse is applied to the platelet line 35.

Alternatively, the detected signal may exceed the low threshold level of the low threshold detector 18 but exceed the low threshold level of the high threshold detector 1.
6, the signal applied to data terminal D of flip-flop FF1 becomes true after being inverted by inverter IN1. Therefore, the next occurrence of the peak detector pulse generated by peak detector 20 and applied to clock terminal CL of flip-flop FF1 will correspond to a platelet pulse applied to platelet pulse line 35. The true output signal appears at output terminal Q of flip-flop FF1. At the same time, ANDGATE AG
1 generates a false signal. This is because the signal on one of the two input lines to AND gate AG1 is false, ie the signal on the line connected to the output of high threshold detector 16 is false. Therefore, when an electrical signal corresponding to detected platelets is generated, only a platelet pulse is generated and no red blood cell pulse is generated for this signal.

In this way, each detected particle that meets the threshold criteria is counted as either a red blood cell or a platelet, depending on which criterion is met. However, in some embodiments it is possible and desirable to generate count increments for both types of particles even if only one type of particle is detected, as shown in Figures 6 and 7. It will become clear below in connection with the description of the figures. In such embodiments, it may be possible and likely that the detection of one or both types of particles within a limited size range will not generate a count increment for either type of particle. It is also desirable to do so.

AND gate AG1 of pulse classification unit 22
is a quadruple two-input positive logic AND gate manufactured by Texas Instruments.
2 such as those available in Model SN408
It is an input positive logic TTL AND gate. Inverters IN1 and IN2 are Hex inverter models manufactured by Texas Instruments.
It's a regular TTL inverter like the one in the SN7404. Flip-flop FF1 is a TTLD type flip-flop unit such as SN7474.

Alternative Embodiment The circuits of FIGS. 2A and 2B show an alternative embodiment of the detection and pulse classification portion of the apparatus of the present invention.

The circuit of FIGS. 2A and 2B is a direct replacement for the portion of the circuit of FIG. 1 between circuit point A' and circuit points E and F. FIG. 2 shows how FIGS. 2A and 2B are combined with each other and with FIG. 3, or alternatively how FIGS. 2A and 2B are combined with each other and with the combination of FIGS. 3 and 4. It shows.

Baseline Return Circuit Baseline return circuit 101 serves the same purpose as previously described in connection with baseline return circuit 14 of FIG. Clamping the combined waveform to approximately 0 volts DC makes it more convenient to discriminate pulses at the basis of their amplitude. As shown in FIG. 2A, the baseline return circuit 101 includes a high-pass filter circuit including a capacitor 102, a PNP transistor 103, and an amplifier 107.
Transistor 103 is oriented in a modified common collector or emitter follower configuration and resistor 10
4,105,106 biased to saturation. The amplifier 107 works as a buffer or isolation device, and the base line return circuit 1
01 output signal BLRO can be applied to a number of other circuits. The input signal to the baseline return circuit 101 is indicated by BLRI.

The special common collector configuration of transistor 103 provides a high input resistance for base current flowing toward its base-collector junction, and a low resistance for base current flowing from the base-collector junction toward capacitor 102. give.
Therefore, the baseline return circuit 101 capacitor and transistor combination provides a DC clamp relative to 0 volts DC. Waveforms WG and WH in Figure 5 are representative input signal BLRI and representative output signal BLRO.
, respectively, showing the clamping operation of the baseline return circuit 101. Also, the baseline return circuit 101
reduces the reliability of the device's detection and counting60
It also acts as a high-pass filter that filters out low frequency signals such as Hz AC signals. Amplifier 107 is a Motorola Model MC1741S, high slew rate, internally compensated operational amplifier manufactured by Monotrola, Inc. of Phoenix, Arizona, USA.

Low Threshold Detector As shown in FIG. 2A, the baseline return circuit 101
The output signal BLRO of is applied to a low threshold detection circuit 111 which includes a voltage comparator 112 and is similar to that described with respect to FIG. 1 and will therefore not be described here. The output signal LLDO of the low threshold detection circuit 111 of FIG. It depends on whether or not it exceeds. The waveform WI in FIG. 5 shows the output signal LLDO in response to the applied signal BLRO of the waveform WH.

Strobe Sequencer Circuit The output signal LLDO of the low threshold detector 111 is applied to a strobe sequencer circuit 121, which consists of a pair of one-shot or monostable multivibrator devices 122 and 123.
These devices are such as the one-shot device found in the Texas Instruments Model 74123 Dual Retriggerable Monostable Multivibrator. Strobe sequencer 121 receives the strobe signal ST used in the circuitry for timing and resetting the pulse generation and detection operations of the embodiment of the invention shown in FIGS. 2A and 2B.
1, ST2, and 2 are generated. Details of how the strobe signals generated in strobe sequencer 121 are used in the circuits of FIGS. 2A and 2B are discussed below. Waveforms WJ and WK in Figure 5
represents strobe signals ST1 and ST2 generated by strobe sequencer 121 in response to signal BLRO of waveform WH.

Peak Hold Circuit The output signal BLRO of the baseline return circuit 101 is also applied to the peak hold circuit 131. The purpose of this peak holding circuit 131 is to generate an output signal EP that follows the BLRO signal as shown in waveform WL in FIG. signal
Two peak levels of signal BLRO (the first peak level corresponds to detected platelets and the second peak level corresponds to subsequently detected red blood cells) until this peak holding circuit is reset by ST2. ).

The transistor 132 of the peak holding circuit 131,
133 and 134 are combined to form a simple differential amplifier, the positive input signal of which is signal BLRO and the negative input signal applied from the source terminal of FET transistor 135 to the base terminal of transistor 133. This is the output signal EP'.
When the voltage level of the output signal BLRO of the baseline return circuit 101 increases, the capacitor 136 of the peak holding circuit 131 is charged via the diode 137. The voltage across capacitor 136 is applied to the gate terminal of FET transistor 135.
FET transistor 135 acts as a unity gain follower. As is clear in Figure 2A,
The source terminal of FET transistor 135 is connected to the positive input terminal of amplifier 138, which is a Motorola Model MC1741S operational amplifier.
Therefore, the peak holding circuit 131 described so far
portion constitutes a means for charging the capacitor 136 in accordance with the input voltage and thereby generating an output signal EP, which signal EP is actually the input signal
It will be clear that as long as BLRO continues to increase it will follow this input signal, but because of the blocking diode 137 it will not be able to follow this input signal BLRO when it decreases. Waveform in Figure 5
WL indicates the tracking and peak holding characteristics of the signal EP.

After this signal EP has been used to count pulses corresponding to red blood cells or possibly platelets, capacitor 136 is discharged and reset to the next pulse corresponding to the next detected particle. Be prepared for the signal. Capacitor 1
36 is connected to the peak holding circuit 1 by the transistor 139 and the strobe sequencer circuit 121.
This is accomplished by the strobe signal ST2 applied to 31. This strobe signal ST2 is approximately 5
It is a positive step waveform with a width of μ seconds (waveform WK in Figure 5).
reference). When this signal is applied to the base circuit of transistor 139, the transistor is turned on,
A coupling portion 142 between each anode of diodes 140 and 141 is grounded. As a result, the positively charged capacitor 136 is discharged through diode 141 to near ground potential, and the peak-holding output signal EP eventually decays to zero volts, as shown by waveform WL in FIG.

As mentioned above, one of the advantageous features of this preferred embodiment of the invention is that the small amplitude pulse corresponding to platelets occurs immediately before the large amplitude pulse corresponding to the subsequently detected red blood cells. This means that they are more likely to be counted as platelets rather than as red blood cells.
This feature of the circuit embodiment shown in FIG. 2A is made possible by the use of a hold signal HOLD applied to the base circuit of transistor 143 of peak hold circuit 131. The manner in which the hold signal HOLD is generated is discussed below in connection with the description of peak detection circuit 161. However, in this respect, the holding signal
HOLD is a positive step waveform (waveform WM in Figure 5).
It is sufficient to note that this is the waveform generated when the signal BLRO begins to decrease and falls some predetermined level below its immediately preceding peak level. This holding signal HOLD
peak hold circuit 1 in order to inhibit further charge buildup on capacitor 136 after a first peak level that satisfies the criteria established for generating this is received by peak hold circuit 131.
31. In this way, the peak hold circuit 131 is made insensitive to signals occurring immediately after the first peak, and the output signal EP of the peak hold circuit 131 is held at the low platelet signal peak and the next detected red blood cell signal. Accidental increases to peaks are avoided.

When a hold signal HOLD, a +5 volt DC step signal, is applied to the base circuit of transistor 143, transistor 143 is turned on. When transistor 143 is turned on, the emitter-base junction of transistor 144 becomes forward biased and transistor 144 is also turned on. When transistor 144 is turned on, resistor 145 is bypassed and the emitter-base junction of transistor 134 is strobed by transistor 144, turning off transistor 134 and preventing further charging of capacitor 136.

Therefore, the output signal EP of the peak holding circuit 131
is a signal that follows the value of the signal BLRO generated by the baseline return circuit 101 until the signal BLRO is held at a constant level when the hold signal HOLD is generated. Signal EP is reset upon application of strobe signal ST2 and is ready for the next signal BLRO.

The degree of pulse coincidence that can still be detected with the initially occurring platelet pulse is determined by the operation of the peak hold circuit 131 in conjunction with the hold signal HOLD, and the signal BLRO before the hold signal HOLD is generated.
depends on the predetermined level that must fall. Typically, approximately 10% below the peak of the red blood cell signal
Platelet signals with peaks occurring μseconds earlier can be distinguished.

Peak Detection Circuit As previously mentioned, the purpose of the peak detection circuit 161 is to generate a +5 volt step signal HOLD each time the level of the output signal BLRO falls by some predetermined amount from its previous maximum value. In the preferred circuit embodiment shown in FIG. 2A, the hold signal HOLD is generated when signal BLRO falls by a maximum of 0.91 BLRO - 50 mV. As shown in FIG. 2A, peak detector 161 is similar to the NSC described above in connection with the threshold detector of FIG.
A voltage comparator 162, Model LM311, is used. The output signal BLRO of the baseline return circuit 101 is applied to the positive input terminal of the voltage comparator 162;
Then, a signal K (EP) which is a fixed multiple K of the output signal EP of the peak holding circuit 131 is sent to the voltage comparator 16.
Applied to the negative terminal of 2. This factor K depends on the values of series resistor 163 and shunt resistor 164, and also on the value of resistor 165 connected to -15 volts DC.

In one form of this embodiment, resistor 163 is approximately 10,000 ohms and resistor 164 is approximately 100,000 ohms.
The resistor 164 is approximately 100,000 Ω, and the resistor 165 is approximately 3,300,000 Ω. resistance 16
If 3,164 and 165 are these values, then K
(EP) is equal to 0.91EP - 50 mV, and the output of voltage comparator 162 is +5 volts as long as the output signal BLRO of baseline return circuit 101 is equal to or greater than K(EP). However, the signal K applied to the negative terminal of voltage comparator 162
When the voltage level of (EP) exceeds the instantaneous level of output signal BLRO, the output signal of voltage comparator 162 changes to 0 volts DC.

The output signal of voltage comparator 162 is applied to one terminal of a three-input NAND gate 166. NAND gate 166 is combined with NAND gate 167 to form a set/reset flip-flop circuit. The output signal of the set/reset flip-flop circuit consisting of NAND gates 166 and 167 is 3.
Applied to one input terminal of input NAND gate 168. The output of this NAND gate 168 is the holding signal HOLD.

As shown in FIG. 2A, the peak detector 161
The second and third input terminals of NAND gate 168 are both coupled to +5 volts DC. The output signal of NAND gate 168 therefore depends on the logic level of the output signal of the set/reset flip-flop consisting of NAND gates 166 and 167. NAND gate 168 inverts the logic level of its input signal so that a hold high signal HOLD is generated when the output signal of the set/reset flip-flop circuit is low.
However, the output signal of the set/reset flip-flop circuit goes low only when the output signal of NAND gate 166 is high and the signals at input terminals 3 and 4 of NAND gate 167 are both high. The signal applied to the input terminal 4 of the NAND gate 167 is the output of the low threshold detector 111, which is the hold signal when the output signal BLRO of the baseline return circuit 101 is less than the 300 mV threshold level.
Prevent HOLD from occurring. Otherwise, the output signal EP of the peak hold circuit 131 may be held at a level corresponding to a noise signal, even though the next pulse actually corresponds to a detected red blood cell. When the next pulse occurs, it is likely to be erroneously counted as a detected platelet.

The third input signal to NAND gate 167 is strobe signal 2, i.e., strobe sequencer 12.
1, and this signal is the logical negation of the strobe signal ST2 already described in connection with the peak hold circuit 131.

The strobe sequencer circuit 121 is a monostable multivibrator (also called a one-shot circuit).
122 and 123. The pulses produced by each one-shot circuit are nominally 5 microseconds wide. Strobe signal ST2 is also used to reset output signal EP of peak hold circuit 131, and strobe signal 2 is the input signal to NAND gate 167 of peak detection circuit 161.

High Threshold Detector Three signals are applied to the circuit shown in FIG. 2B: peak hold output signal EP, strobe signal ST1, and strobe signal ST2. Signal EP is applied to the positive input of high threshold detector 171, which includes voltage comparator 172 and operates as previously described in connection with FIG. Negative terminal 3 of comparator 172
The voltage applied to is the reference potential with which the signal EP is compared. This reference potential is nominally about 3.5 volts DC, which corresponds to the threshold level for the typical smallest red blood cell size.

Pulse Classification Unit The output signal of the high threshold detector 171 is passed through the D-type flip-flop of the pulse classification unit 181.
Applied to data terminal D of flop 182. A strobe signal ST1 is applied to the clock terminal CL of this flip-flop 182. The output terminal Q of flip-flop unit 182 is connected to one input terminal of double-input NAND gate 183, and
The output terminal of the flop unit 182 is 2.
It is connected to one input terminal of the multiple input NAND gate 184. Strobe signal ST2 is connected to the other input terminal of each of these NAND gates 183 and 184.

If the signal EP produced at the output of the peak hold circuit 131 shown in FIG. 2A exceeds the threshold level established for the smallest sized red blood cells, then the high threshold shown in FIG. The output signal of the hold detector 171 is approximately +5 volts DC. As a result, a positive strobe signal
When ST1 subsequently occurs, output terminals Q and provide positive and negative, ie high and low, signals to NAND gates 183 and 184, respectively. However, since strobe signal ST2 is normally 0 volts DC, the output terminals of NAND gates 183 and 184 will generate a high logic level signal until this strobe signal ST2 goes positive, causing signal ST
When 2 becomes positive, the output signal of NAND gate 183 changes from a high level voltage to a low level voltage, and the negative going edge of the signal on pulse line 33 is counted as a red blood cell pulse.

On the other hand, the signal level of the output signal EP of the peak holding circuit 131 is 3.5, which corresponds to the detected platelets.
volts, the output signal of high threshold detector 171 is approximately 0 volts DC, and strobe signal ST1 is applied to the clock terminal of D-type flip-flop 182.
When applied to CL, flip-flop 18
2 output terminals Q and are respectively negative and positive,
That is, it generates low and high level voltages. after that,
When the high level strobe signal ST2 is received by the pulse classification unit 181, the output of the NAND gate 184 on the pulse line 35 changes from positive to negative, i.e. from high level to low level (fifth
(See waveform WN in the figure). Platelet pulses are therefore counted.

The pulse classification unit in Figure 1 generated positive-going pulses for detected platelets and red blood cells, whereas the pulse classification unit in Figure 2 generated negative-going pulses for detected red blood cells and platelets. Please note that. However, the third
As will be understood from the detailed description of the counting circuit of the present invention in conjunction with FIG. Count its return edge or, in the case of a negative pulse, its starting edge. It is not intended that the scope of the invention be limited to any particular voltage polarity or magnitude.

Counting Circuit FIG. 3 shows a detailed diagram of the counting section of the present invention, which includes a platelet pulse counter 24, a platelet count display unit 26, a red blood cell pulse counter 28, and a thumb-operated wheel switch arrangement 3.
It is equipped with 0.

Pulses corresponding to detected platelets are sent from pulse classification unit 22 in FIG. 1 or unit 181 in FIG.
2 enabling terminal pressures are applied. This binary coded decimal counter 32 generates one positive pulse at its output terminal Q4 for each platelet pulse sent via line 35. Motorola's Model MC14518cp has two binary coded decimal up counters, each of which
2 is suitable for use in the present invention.

The output of the binary coded decimal counter 32 is applied to line 37 which is the input line to a three digit binary coded decimal counter 34. This 3-digit binary coded decimal counter 34 is connected to its clock terminal C.
gives a three-digit decimal count of the number of pulses applied to Counter 34 also includes a multiplexer (not shown) and a scanning oscillator (not shown) that time-multiplex each of the three 1-digit counters to terminal Q.
The output signals of 0 to Q3 are each sequenced and these signals are combined to output 10 of the first, second or third digit at a time.
Be able to represent decimal numbers. The digit select signals available at terminals 1, 2, and 3 are also sequenced by the scanning oscillator to operate the appropriate decimal indicators synchronously with respect to the decimal digits generated by the three-digit counter 34. . Motorola Model
The MC14553CP, 3-digit BCD counter is one example of such a counter suitable for use with the present invention. A capacitor C _f connected between terminals 3 and 4 of the three-digit binary coded decimal counter 34 determines the scanning frequency used to time multiplex the output of the counter 34. For purposes of this invention, an acceptable value for capacitor C _f is 1000 pf.

The output signals of the three-digit binary encoded decimal counter 34 are applied to lines 39, 41, 43, and 45, respectively, and sent to a decoder 36. Decoder 36 is 4
A seven segment display device 40 for receiving a bit binary coded decimal signal and decoding such signal;
42 and 44. One such decoder suitable for use with the present invention is the Texas Instruments Model SN7447A, BCD-7 Segment Decoder/Driver. The decoder 36 is configured so that each of the seven output signals from the decoder unit 36 is current limited by a resistor of an appropriate value.
The output signal of is applied to the resistor network. Approximately 75 each
A Sprague type 914C-SR resistor network having a set of seven resistors of Ω is a suitable resistor network for use in the present invention.

The output lines of resistive network 38 are connected to three decimal indicators 40, 42, 44, respectively. There are a number of methods suitable for use in the present invention for the purposes set forth herein.
Although decimal indicators are available, the preferred embodiment of the present invention uses three common anode LED displays, each having characters approximately 0.3 inches high. Model by Heuretsu Pats Card Company
A 5082-7730 solid state seven segment indicator is an example of a decimal indicator suitable for use with the present invention.

Each of the indicators 40, 42 and 44 is set to +5 by transistor switches 46, 48 and 50, respectively.
It has a single anode connected to a volt DC. Only one switch is closed at a time by a digit select signal generated at terminals 1, 2, and 3 by a three-digit binary coded decimal counter 34. If the frequency of the scanning oscillator of the 3-digit binary coded decimal counter 34 is high enough, for example greater than 100 Hz, there will be no visible flickering due to visual afterimages, and the decimal indicator will display continuously. It looks like it's emitting light.

The three-digit binary coded decimal counter 34 may be replaced with three or more separate counters and also indicates an overflow condition or when a very high particle count exceeds the maximum number that can be displayed. It will be apparent to those skilled in the electronic arts that circuit means may be provided to stop the counting.

Platelet pulse counter 24 and platelet pulse count display 26 thus constitute a counting and display device that generates a count of platelet pulses applied from pulse classification unit 22 and displays the three upper digits of this count. is clear;
The above digits are thousands of platelets per microliter of the blood sample being tested, as will be seen below. As the platelet pulse count progresses, the displayed digits are divided into 3 digits.
It keeps changing based on the output signal of the decimal counter.
In most cases, the platelet count display changes slowly enough to be visually observable during the counting process. The rhythm established by this count is a useful indication of the uniformity of the sample flow through the transducer.

Once the counting process is complete, the final platelet count is displayed until power is removed or the counter is reset as described below.

The red blood cell pulse is detected by the pulse classification unit 22 or 1.
81 via line 33 to red blood cell pulse counter 28 and applied to binary coded decimal counter 52. This counter 52 is a binary coded decimal counter 32 used in the platelet pulse counter 24.
This is the same type of counter as . An input pulse is applied to the terminal pressure or enabling terminal of counter 52, and an output signal is obtained at terminal Q4. The output signal of counter 52 is a pulse generated once every ten input pulses are counted. The output signal of binary coded decimal counter 52 is applied to line 53, which is connected to the enable terminal pressure of binary coded decimal counter 54. This counter 54 is a binary coded decimal counter 52
This is the same type of counter. The output signal of this counter 54 is available at terminal Q4 and is a pulse generated once every ten input pulses are counted. Therefore, counter 54 generates one pulse for every 100 red blood cell pulses generated by pulse classification unit 22. Therefore, the binary coded decimal counters 52 and 54 have a fixed two-digit value.
A decimal (ie 100) divider or pre-conversion device 51 is constructed. An alternative pre-scaling circuit which provides a degree of adjustability and which directly replaces this pre-scaling circuit 51 is shown in FIG. 4 and will be described below.

In this regard, it should be recalled that the purpose of counting red blood cells together with platelets is to arrive at a count of red blood cells corresponding to the already known or assumed red blood cell concentration of the blood sample being examined. . When the red blood cell count reaches a number corresponding to this known or assumed concentration, the counting process carried out by both the red blood cell pulse counter 28 and the platelet pulse counter 24 is stopped and the display is then displayed. It is desirable that the number of platelets taken corresponds to the platelet concentration of the blood sample being tested.

As mentioned above, it is possible to stop only the platelet pulse counting process when the red blood cell pulse counter reaches the preset count, and to only count platelets without stopping any counting process when the red blood cell pulse counter reaches the preset count. Each recording is also within the scope of the present invention.

Before starting the counting operation, the three upper digits of the previously known red blood cell count concentration are set on the binary coded decimal thumb-operated wheel switches 65, 75, 85 constituting the thumb-operated wheel switch array 30; recorded and retained.
During the counting operation, the output signal of the binary coded decimal counter 54 is transmitted to the three binary coded decimal counters 56, 66, 76.
are applied to a series combination of . These three counters are connected in series so that each produces a count corresponding to a decimal number of one of the three high order digits of the red blood cell count.

Binary coded decimal counters 56, 66 and 76 are each of the same type as binary coded decimal counter 52. Each thumb-operated wheel switch of the thumb-operated wheel switch array 30 may be configured, for example, by Digitran.
Model 28015 is a binary coded decimal thumb-operated switch. Each binary coded decimal counter 56, 66, 76
are the output terminals Q1, Q corresponding to the corresponding binary digit fields 1, 2, 4 and 8 of the decimal number, respectively.
2, Q3 and Q4.
Each such output signal is applied via a diode to the appropriate binary column terminal (ie, 1, 2, 4, or 8) of each thumb-operated wheel switch 65, 75, 85. The output signal at terminal Q4 of binary coded decimal counter 56 is applied to the enable terminal E of binary coded decimal counter 66, and the output signal at terminal Q4 of binary coded decimal counter 66 is applied to the enable terminal E of binary coded decimal counter 76. voltage is applied to terminal E. As a result, the binary coded decimal counter 56 is
The binary coded decimal counter 66 generates one pulse at its terminal Q4 every time 1000 red blood cell pulses are counted, and the binary coded decimal counter 66 generates one pulse at its terminal Q4 every time 10000 red blood cell pulses are counted.
generates one pulse. Therefore, for example, the first
The actual count corresponding to the digit 488 shown displayed on thumb wheel switch array 30 is 48,800 red blood cells. A red blood cell count of 48,800 effectively corresponds to 0.01 of a blood sample containing 4.88 million red blood cells per unit volume.

As mentioned above in connection with Figure 3, the red blood cell count is
Prescaled by a factor of 100 and platelet counts prescaled by a factor of 10. Therefore,
Thumb operated wheel switch array 30 has a value of 488
, the actual number of red blood cell pulses on the red blood cell pulse line 33 corresponding to such a count is 48,800. Thus, for a diluted sample having a concentration of 48,800 red blood cells per microliter, one milliliter of this diluted sample is flowed through the sensing region of the transducer to produce such counts. At the same time, the number of platelet pulses occurring on platelet pulse line 35 is approximately 3040 during this same interval. As is clear, 1/of the original concentration
3040 platelets in a microliter volume of whole blood diluted to 100 corresponds to a concentration of 304,000 platelets per microliter of undiluted whole blood. Therefore, prescaling the platelet pulse count by 10 yields a count reading of 304, which is then expressed as thousands of platelet counts per microliter of whole blood.

Each of the binary coded decimal counters 56, 66, and 76 produces approximately 0 volts DC at terminals Q1, Q2, Q3, and Q4 for binary 0s and at these terminals for binary 1s. Each generates approximately +5 volts of DC. Binary coded decimal counter 5
As long as the binary count produced by each of 6, 66, or 76 is less than the decimal count set in each of its corresponding thumb-operated wheel switches 65, 75, 85, common line 90 is connected to diode 5.
8, 60, 62, 64, 68, 70, 72, 7
4, 78, 80, 82 and 84 remain electrically connected to ground potential. However, binary coded decimal counters 56, 66,
The binary counts generated by each of 76 are transmitted to the corresponding thumb operated wheel switch 65,
When it becomes equal to each setting of 85,
None of the 12 possible paths through the appropriate diodes are connected to ground potential G. At this time, no current flows through the pull-up resistor R _p and the voltage on line 90 increases to +5 volts DC, generating a true end-of-count signal STOP.

As shown in FIG. 3, the counting end signal STOP is applied to the clock terminal C of the binary coded decimal counter 52 of the red blood cell pulse counter 28 and the clock terminal C of the binary coded decimal counter 32 of the platelet pulse counter 24. is applied to True count end signal
STOP inhibits further changes in these two binary coded decimal counters, so both counting processes are immediately terminated.

The reset line R/S can be used to reset all binary coded decimal counters before starting a new test. +5 reset line
When connected to a volt source, all counters are reset to zero counts and ready for a new test. The reset line R/S is connected to the source by a conveniently located momentary spring loaded switch (not shown) to make it convenient for the operator to reset these counters at appropriate times. connected to.

As mentioned above, certain pre-scalators 51 shown in FIG. 3 may be replaced by pre-scalers that provide some degree of adjustability. Such an adjustable preconversion device 251 is shown generally in FIG. Replacing the constant pre-scalator 51 with the adjustable pre-scaler 251 allows the red blood cell count to be divided by any integer within the range of 100 to 109.

Dividing by an integer other than 100 in a red blood cell counter biases the red blood cell count, thereby causing higher platelet concentrations and correlating more closely with other types of platelet counters that normally indicate such high concentrations. Sometimes desired for.

Details of the adjustable preconversion device will be explained below with reference to FIG. As shown in Figure 4,
The adjustable pre-conversion device 251 has two binaries.
It includes decimal counters 252 and 254, which are connected to a predetermined conversion device 5 of FIG.
1 binary coded decimal counters 52 and 54 in all respects. Red blood cell pulse line 33 is 2
Connected to enable terminal E of evolved decimal (BCD) counter 252. This BCD counter 25
2 generates a pulse at its terminal Q4. Connection line 253 transfers the output pulses of BCD counter 252 to enabling terminal E of BCD counter 254. BCD counter 254 generates a pulse at its output terminal Q4 every time the tenth input pulse is applied to its enable terminal E. Therefore, BCD counters 252 and 254
operates in exactly the same way as the two counters 52 and 54 of the constant prescaling device 51 of FIG. However, unlike the preconversion device 51 of FIG. 33 and is not applied to line 55 until it is received by this preconversion device 251.

The output signal at terminal Q4 of BCD counter 254 is applied by conductors 268, 269, and 270 to an inverter in the form of a NOR gate 260. The output signal of NOR gate 260 is connected to D by conductor 271.
applied to the clock terminal CL of the flip-flop 258. This D-type flip-flop 258 is, for example, Motorola Model MC4013.
This is one half of a dual D flip-flop circuit. As is clear from FIG. 4, data terminal D of flip-flop 258 is always connected to +5V DC. As a result, when this flip-flop is clocked by a pulse applied to its clock terminal CL, its output terminal Q will be positive, or true, and its output terminal will be negative, or false. Output terminal Q of flip-flop 258 and clock terminal of BCD counters 252 and 256
Each is connected to CL. A positive signal applied to the clock terminal of counter 252 inhibits further changes in this counter and
A negative signal applied to the clock terminal of 6 enables this counter to begin counting the pulses sent to its input terminal E. Therefore, in practice, the D-type flip-flop 258 converts the red blood cell pulses sent via the pulse line 33 to the preconversion device 251 into the BCD counter 252 or 25.
This is a judgment gate that allows one of 6 to be counted. It will be appreciated that switching the counting of pulses on line 33 from BCD counter 252 to 256 occurs when 100 pulses have been counted by BCD counter 252.
Output terminals Q1, Q of BCD counter 256
2, Q3 and Q4 are BCD counters 56, 6
Diodes 263-26, respectively, are connected to respective thumb-operated wheel switches 65, 75, 85, as previously described with respect to FIG.
6 to the thumb operated switch 257.

The common terminal C of the thumb operated wheel switch 257 remains at ground potential or 0 volts DC until the BCD counter 256 reaches a count corresponding to the count set on this switch 257. Since the common terminal C of thumb-operated wheel switch 257 is connected by conductor 282 to the D terminal of D-type flip-flop 259, the state of this flip-flop 259 also remains fixed and its output terminal is There is approximately 5 volts DC, corresponding to the true condition, and output terminal Q is O volts DC, corresponding to the false condition. The clock terminal CL of the D-type flip-flop 259 is connected to the pulse line 33 by a connection 275, and thus this flip-flop
Flop 259 is clocked each time it receives a pulse on line 33.

The D terminal of the D flip-flop 259 is also connected to the Q terminal of the D flip-flop 258 via a resistor 267 by a connection 273. Resistor 267 acts as a pull-up resistor similar to resistor R _p of FIG. Terminal Q of flip-flop 258 serves as a 5 volt DC source to connect terminal D of flip-flop 258 to +5 volts DC when the common terminal of master operated wheel switch 257 is disconnected from ground potential. Thumb operated wheel switch 2
57 common terminal C is disconnected from ground potential when BCD counter 256 reaches a count equal to the count set in this switch 257. In response, the voltage at terminal D of flip-flop 259 increases to the voltage at terminal Q of flip-flop 258, approximately +5 volts DC. As a result, the D-type flip-flop 259
The output terminals Q and of change their state upon the occurrence of the next red blood cell pulse received via line 33. At this time, the output terminal Q of the flip-flop 259 is positive, that is, about +5 d.c.
volts, and the output terminal becomes negative, that is, about 0 volts DC. The negative going signal of the output terminal of flip-flop 259 is then applied to line 55, which is connected to enable terminal E of 100x BCD counter 56 as shown in FIG.

Therefore, this adjustable preconversion device 251
Therefore, the number of pulses received via line 33 is
Each time the number equals 100 plus the number set on thumb-operated wheel switch 257, line 5
It can be seen that a pulse is applied at 5.

Output terminal Q of D-type flip-flop 259 is connected to one of NOR gates 261 by conductor 277.
connected to one input. The output of NOR gate 261 is connected by conductor 278 to both input terminals of NOR gate 262. Noah Gate 262
The output of is via connections 279, 280, 281
They are connected to the reset terminals of a BCD counter 256 and a D-type flip-flop 258, respectively. Therefore, when the signal at output terminal Q of D-type flip-flop 259 becomes positive, a positive signal is applied to each reset terminal of BCD counter 256 and D-type flip-flop 258. As a result, BCD counter 256 is reset to a count of 0 and flip-flop 258 is reset so that the signal at its output terminal Q is false, or approximately 0 volts, and the signal at its output terminal is true, or approximately 5 volts DC. Ru. accordingly
When BCD counter 256 is reset to 0,
This counter 256 is connected to its clock terminal C.
BCD counter 252 is also disabled by setting the voltage at its clock terminal C to positive, and BCD counter 252 is enabled by setting the voltage at its clock terminal C to negative, whereby BCD counters 252 and 254 are disabled. The next 100 pulses received via pulse line 33 are ready to be counted.

To illustrate the operation of adjustable preconversion device 251, thumb-operated wheel switch 257 is set to the number 5 so that a negative going pulse is placed on line 55 for every 105 pulses received on line 33. Assume that the voltage is applied. first
While 100 pulses are received, BCD counter 2
52 and 254 are enabled and BCD counter 256 is disabled. When the 100th pulse is received on line 33, BCD counter 254 produces a positive output signal at its terminal Q4. In response, D-type flip-flop 258 is clocked and its output terminal Q
is set positive, its output terminal is set negative, and BCD counter 252 is disabled and BCD counter 256 is enabled. This BCD counter 256 then takes over and advances the count by further pulses received via line 33. 105th pulse, i.e. BCD
During the enabled state of counter 256, this counter 2
When the fifth pulse, received by 56, is received via line 33, BCD counter 256 produces an output corresponding to a count of 5 on each of its output terminals Q1 through Q4, so that this count is This is preset in switch 257. As a result, the common terminal C of the thumb-operated wheel switch is disconnected from ground potential, or 0 volts DC, and the data terminal D to the D-type flip-flop 259 reaches the potential of the output terminal Q of flip-flop 258, or about 5 volts DC. urge Thereafter, when a clock pulse is received by flip-flop 259, the terminal of flip-flop 259 applies a negative going signal to line 55, which signal is counted by BCD red blood cell counter 56 (FIG. 3). ).

If division by 100 is desired, as was done in the pre-conversion device 51 shown in FIG.
occurs immediately when thumb-operated wheel switch 257 is set to 0 and thus enabled.
It is observed that the zero state of BCD counter 256 causes the data terminal of flip-flop 259 to reach the +5 volt DC potential of output terminal Q of flip-flop 258.

The reset signal on reset line R/S and the end-of-count signal STOP on line 90 perform the same functions as described above with respect to FIG.

Coincidence effects and other sources of error If the flow rate of red blood cells through the particle transducer is very low, either very few red blood cells will be counted or the test will take a long time to complete. That is clear. In either case, a low total count will result in statistical sampling errors since the red blood cells are not evenly spaced along the axis of the channel. On the other hand, if the particle concentration is very high, some particles will flow into the transducer close to each other, thus producing superimposed pulses. In the present invention, the total counts for each type of particle are made large enough to avoid significant statistical errors in small ranges, and below the value corresponding to the maximum interference between pulses due to approach effects. The flow rate is set to . Furthermore,
In the present invention, pulses having amplitudes of different ranges are counted and the ratio of the counts determined to substantially reduce the errors that are likely to occur due to such interference.

When two types of particles are counted, the proportion of error in both counts is approximately equal, so that the error in counting one type of particle is due to the error in counting the other type of particle. be compensated for. In the present invention, such compensation is achieved by determining the ratio of counts during equal time periods, or more simply, by using the occurrence of a given count of one particle, This is achieved by automatically determining the time at which the other particle is counted. In either case, knowing the concentration of one type of particle, such as red blood cells, helps determine the concentration of another type of particle, such as platelets. It is to be understood that concentration values are expressed in terms of particles per microliter or hematocrit or volume fraction.

It should be appreciated that due to coincidence interference, the number of pulses detected is less than the number of particles flowing to the transducer. The difference between the number of particles flowing through the transducer and the number of pulses counted is often referred to as coincidence loss.

The rate at which the pulses are generated is approximately given by:

n=N(1-CN) where n is the number of pulses generated per second, N is the number of particles flowing into the transducer per second, C is the coefficient of coincidence loss, and The "coincidence loss" in unit time is given by the product of ^CN2 .

The total number of particles detected during the time interval T is nT. The statistical error, ie the difference in counts detected during a short time interval T compared to a long time, can be reduced by increasing T.

It can be shown that in general the counting rate has a maximum theoretical value n=1/4C.

For a particle transducer with a length of 78μ and a diameter of 55μ, a series of experiments determined the value of the coincidence loss coefficient C to be C = 43.6μsec/red blood cell, where the flow rate is equal to the number of red blood cells flowing through the transducer. The transition time was set to be 66 μs.

A method for measuring the match loss coefficient was developed in 1965.
October 21-22, San Francisco, California, Proceedings of the 5th Counter Counter
“Calibration, Coincidence Correction and
Interpretation of Counter Counter Data” and October 1966, Review of
Scientific Instruments Vol. 37, No. 10, pp. 1416-
“Improved” by LHPrincen of 1418
Determination of Calibration and Coincidence
“Correction Constants for Counter Counters”
It is described in.

The corresponding maximum theoretical counting rate is n _nax = 5733
It is calculated as red blood cells/second.

When operating at a counting rate of 3000 pulses/sec, slightly above half this theoretical maximum, the actual number of red blood cells flowing through the transducer is 3500;
This represents a match loss error of 14.3%.

As a first approximation, since the concentration of platelets is small compared to the concentration of red blood cells, the error due to coincidence of platelets flowing into the transducer at the same time as red blood cells will be about the same, ie, 14%. By electronically automatically counting pulses due to red blood cells and pulses due to platelets electronically, and automatically turning off counting of platelet pulses when a predetermined red blood cell count is reached as described above, at least In theory, the two errors will completely compensate each other and indicate the correct value of the platelet count. Even if the red blood cell concentration is high and the red blood cell flow rate is
Even with as few as 11,000 particles, substantially correct results can be obtained with substantially complete compensation for matching errors. The actual results are not as good. In particular, in the form of the invention shown in FIGS. 2A and 2B, pulses are more likely to be counted by the red blood cell counter as platelets flow through the transducer in substantially unison with red blood cells. This creates a masking effect. In fact, in this example of the invention, when the red blood cell count rate is 3000 red blood cells/second, the platelet count loss for approximately 90% or more of the test series is 3% due to various causes including coincidence effects. It was between 7% and 7%.

The reason for this is that red blood cells are counted more frequently because some platelet pulses are lost during the sharp rise of the red blood cell pulse when a near match occurs.

By taking the ratio of the two pulse numbers, the error due to coincidence alone is substantially reduced if the pulse numbers are about 20% to 75% of the maximum. In the best embodiment of the invention, this ratio is automatically calculated electronically. If systematic errors appear, the red blood cell pulse preconversion device 251 (fourth
The readings are automatically corrected by setting the appropriate platelet count (Figure) to the appropriate value so that the correct platelet count can be read directly from the meter.

Under certain conditions, all matches are red blood cell-like 1
It is read as a type of particle. For example, this is
This occurs in the first embodiment of the invention where no precautions were taken to control the counting by the first of two superimposed pulses. In such a case, all the effects of the present invention cannot be obtained. On the other hand, when the first pulse detection and signal holding features are used as in the second embodiment of the present invention, the effects of the present invention can be obtained over a wide range of areas.

In most cases, the size of red blood cells lies outside of the size range of platelets and can also be reacted with this. However, there are cases where the size range of red blood cells and the size range of platelets overlap at least to some extent. It is generally known that there are very few platelets with a size of 27.5μ3 ^or more and very few red blood cells with a size of 27.5μ3 ^or less. Therefore, measure the concentration of red blood cells of ^27.5μ3 or higher and treat them as if they were red blood cells, even if some of them are platelets and some of them are white blood cells. is common. To this end, the threshold of the high threshold detector 16 is set at a red blood cell size of ^27.5μ3 in order to activate the red blood cell pulse counter using pulses generated by particles larger than ^27.5μ3 . is set to the voltage corresponding to the voltage. In this case, even if particles other than red blood cells are counted, the pulse counter 28
It is interesting to note that generates a signal that corresponds to the original volume of the blood sample.

If a "red blood cell" count is derived from a machine such as a Coulter S machine and all particles have a volume greater than or equal to the reference size being counted, then this count can be applied to a specific known volume of blood. Please note that it is compatible with Thus, when the apparatus of the present invention is similarly set to count all such particles, the pulse counter 28 will correctly indicate when the same volume of blood has flowed through the particle sensing transducer 10. Therefore, the corresponding indication generated by platelet pulse counter 24 indicates the number of platelets of size 27.5μ3 ^or less within this same volume. Therefore, in this regard, it should be noted that the platelet concentration is determined correctly, except for some errors caused by a small proportion of particles having a larger volume than the reference amount.

Variations of the Invention From the foregoing description of the invention, it is clear that the threshold voltage corresponds to a particular size of particles when a blood sample flows through a particular transducer. Therefore, the potentiometer can be set at various positions to generate pulses corresponding to two predetermined size ranges of particles.

When using red blood cell counts determined by other machines, the threshold can be set at a point such that the red blood cell counts generated by the device of the present invention correspond to the counts generated by the other machines mentioned above. desirable.
This applies even if other machines incorrectly count some particles other than red blood cells and treat them as if they were red blood cells.
Such errors occur in virtually all so-called red blood cell counters currently on the market, which count all particles having a volume of 27.5 microns ^or more. Therefore, in such machines, white blood cells are counted as red blood cells because they are larger than red blood cells, and the count also includes some of the larger platelets. When counting platelets according to the present invention, the threshold separating red blood cell pulses from platelet pulses is typically set to a value corresponding to about ^27.5μ3 . The resulting error in platelet concentration is small.

It has been found that there are a significant number of blood samples that have a platelet distribution such that a significant number of platelets have a size of approximately ^30μ3 . For this and other reasons, it is desirable to provide an arrangement such that the low threshold used to count red blood cells is below the high threshold at which the pulses being counted involve platelets. It can be done. Two simple circuits for achieving this result are shown in FIGS. 6 and 7, respectively.

In the configuration of FIG. 6, the high threshold detector 16 of FIG. 1 is replaced with two threshold detectors 16' and 16''. Hold detector 18 is referred to as a first low threshold detector. Threshold detector 16'' is
It is called a high threshold detector because it determines the high threshold detection level of pulses that are counted as platelets. On the other hand, the threshold detector 16' is referred to as a second low threshold detector because it detects the low threshold detection level of pulses counted as red blood cells. This low threshold detector 1
The output of the high threshold detector 16'' is connected to the AND gate AG1, and the output of the high threshold detector 16'' is connected to the flip-flop FFL via the inverter IN1.
is connected to terminal D of The two threshold detectors 16' and 16'' are set at different levels, one above the other.

In this configuration, the second low threshold detector 16' establishes a minimum cutoff point for pulses that are treated as representing red blood cells. A pulse having an amplitude above this threshold as determined by the second lower threshold detector 16' generates a red blood cell signal on line 33. The maximum cutoff point for red blood cells is a function of circuit element constraints.

In the configuration of FIG. 6, if the detected pulse has an amplitude above the threshold set in the high threshold detector 16'',
The pulse appearing at circuit point N is a flip-flop
Ban FF1. Therefore, if the pulse has an amplitude in the range between the low level established by the first low threshold detector 18 and the level established by the high threshold detector 16'' , a pulse appears on line 35, otherwise it would not appear.

In the configuration of FIG. 6, if the threshold established in detector 16'' is above the threshold set in the second lower threshold detector 16', the pulse will be detected on line 33.
and 35, so each pulse within that range is counted as both red blood cells and platelets, even though the pulses normally represent only one of red blood cells and platelets. This mode of operation is particularly useful when the machine originally used to count red blood cells in the sample incorrectly counts large platelets as if they were red blood cells. As explained in more detail below, the second low threshold detector 16' generates a number of pulses on line 33 and 35 on line 35 corresponding to the red blood cell counts made even in error. is set at a point that allows the number of pulses obtained to correspond to the concentration of platelets in the sample. Specifically, with this configuration, pulses are generated on line 33 corresponding to those generated in the machine that incorrectly counted such large platelets as red blood cells.

On the other hand, if the threshold established for the second low detector 16' is above the threshold set for the high threshold detector 16'', no pulse will be generated on line 33 or 35. Thus, pulses that are within the gap between the upper limit of the platelet pulse amplitude range and the lower limit of the red blood cell pulse amplitude range are not counted at all.

For example, platelet threshold detection circuit 1
8 and 16″ are ^2.0 μ to detect platelets.
The second lower threshold at which particles are detected by detector 16' is set at a voltage corresponding to a particle having a volume of ^30.0μ3 , while the red blood cells and 27.5μ to detect large platelets counted as
The voltage is set to correspond to a particle with a volume of ³ . In either case, the counting units 24, 28 and their associated circuitry operate to display the count of the platelet pulse counter 24, which approximates the concentration of platelets, as described above.

The structure of FIG. 7 is similar to that shown in FIG. 2B. In this case, the high threshold detector 171 of FIG. 2B is replaced by two threshold detectors 171' and 171'' identical to the threshold detectors 16' and 16'' of FIG. and the pulse classification unit 181 of FIG. 2B is divided into two pulse classification units 1.
81' and 181''. In this case, the flip-flop 182 of FIG. 2B is replaced by two flip-flops 182' and 181'', respectively.
It has been replaced with 2″.And Gate 183
One branch of AND gate 184 is connected to output terminal Q of flip-flop 182', and one branch of AND gate 184 is connected to output terminal Q of flip-flop 182'.
The other input branches of the two AND gates 183 and 184 are each connected to receive the strobe signal ST2. The data terminal D of the flip-flop 182' is connected to the output terminal of the second AND gate ST2. and the data terminal D of the flip-flop 182'' is connected to the output terminal of the high threshold detector 171''. In this case, in the case of FIG. As shown in Figure 2, the height of the detected pulse is the second
A pulse is generated in line 33 to be counted as a red blood cell pulse if the threshold determined by the low threshold detector 171' is exceeded, and the pulse height is 2A) and the high threshold detector 171'', a pulse is generated to be counted as a platelet. In this case, as in FIG. If the two ranges overlap, the same pulse will be counted as a red blood cell pulse and a platelet pulse. Similarly, pulses whose width is within the gap between the two ranges will not be counted at all.

A typical threshold setting for a high threshold level or upper limit for detecting platelets corresponds to a voltage V2 of 3 volts for high threshold detector 171''.
A typical threshold setting for the second low threshold level or lower limit for detecting red blood cells is a voltage V1 of 2.75 volts for the second low threshold detector 171'.
corresponds to 3.0 volt setting is 30μ ³
and a setting of 2.75 volts corresponds to a volume of ^27.5μ3 .

CONCLUSION The present invention uses electronic particle counting to simultaneously count red blood cells and platelets and automatically detect and display the platelet concentration of a blood sample, and which can be previously measured or It should now be clear that the count is controlled by the detected red blood cell concentration.

It will also be appreciated that in the present invention, even if the volume of the diluted sample is not measured, the platelet concentration of this diluted blood sample is determined and thus represents the entire blood supply of the test subject. actually,
The accuracy of measuring platelet concentration according to the invention does not depend on accurately knowing the volume being measured, nor does it depend on the accuracy of the dilution with which the sample is prepared.

Additionally, it will be appreciated that a number of embodiments have been disclosed herein, including preferred embodiments of apparatus specifically configured to carry out the method. This device uses one of a number of known particle sensing transducers that generate an electrical signal whose amplitude depends on the size of the particle being sensed. This device distinguishes between the electrical signals generated by detecting platelets and the electrical signals generated by detecting red blood cells, and supports separate lines, ie, a line that works to count red blood cells and a line that counts platelets. It serves as a means for generating pulses.

Furthermore, the device of the invention comprises counting means for separately counting such pulses until the number of red blood cells counted is equal to a preset value, and for performing separate manual or mental calculations. It also includes means for generating a count end signal for automatically terminating the counting process when the displayed platelet pulse count directly corresponds to the platelet concentration of the test sample.

Although in describing the invention, reference has been made to specific illustrative embodiments thereof, numerous changes and modifications will be apparent to those skilled in the art without departing from the scope of the invention. For example, the method of the invention may be achieved using folding arrangements other than the specific embodiments described herein, i.e. using a particle sorting device in which coincidence counts can be generated separately and stopped by manual means. It should now be clear from the teachings herein that it may be used. As yet another example, a model of multi-channel analyzer with pulse height analysis, pulse counting and count display capabilities, such as the Nuclear of Schuyerburg, Illinois, USA.
It will also be apparent that the present invention can be practiced in connection with the Model ND100 multi-channel analyzer manufactured by Data Inc.

A 1974 book describing the Nuclear Data ND100 multichannel analyzer, and “IM88− for control operation of the ND100 multichannel analyzer.
0551-00, Operator's Instruction Manual,” was published by Nuclear Data Inc. and is incorporated herein by reference. These two publications ensure that the ND100 It will be clear to those skilled in the art how to implement the invention in connection with a multi-channel analyzer.

The use of this particular multichannel analyzer is not as satisfactory as the apparatus detailed above. This is because the multichannel analyzer has no means for stopping the counting process when the red blood cell count corresponds to a number other than the specific count for which it was calibrated. Therefore, for example,
When it is desired to stop the counting process at a point corresponding to 4.88 million red blood cells per microliter, the closest stopping point that can be obtained with this particular multichannel device is 5 million red blood cells per microliter. Therefore, when using such a device, further correction operations must be performed if accurate counting is desired. However, it is possible to use a lever device with an arrangement in which the counting interval is measured in time.

In view of the foregoing description, the present invention is primarily concerned with a method and apparatus for measuring counts per unit volume, i.e. the density or concentration of particles in a liquid sample of variable volume, and the scope of the invention is It will now be apparent that there are many variations of the elements of the apparatus and steps of the method that may occur, and the invention is not limited to the specific embodiments described.

The type of circuit used to sense, discriminate, classify and count is responsive to electrical signals generated by this same device and applied to the counting device as an indication of the reference particle concentration; It will also be apparent that modifications can be made to eliminate the need for manual setting of reference concentrations.

Still further variations in the steps and/or elements of the invention described herein include steps and means for determining the density or concentration of a plurality of different types of particles, including those having unknown densities or concentrations. Also included. Yet another modification is to add a known concentration and volume of synthetic or non-synthetic particles to a known volume of sample fluid to synthetically produce a reference particle of appropriate concentration, which is relative to the particle under test. Further steps and means adapted for measuring concentration are used.

It is therefore to be understood that the invention is intended to cover all such changes and modifications as may justly and properly fall within the scope of the appended claims.

As explained in the introduction to this specification, the broad concept of the invention applies to concentrations or densities measured and expressed in other ways than counts per unit volume, particularly in terms of hematocrit in percent volume. You can also. In many laboratories, the hematocrit of a blood sample is established by the well-known centrifugation process. Therefore, hematocrit, rather than red blood cell count per unit volume, can be used many times more conveniently as a characteristic of a particular sample of blood. In other words, it is more convenient to enter hematocrit data, rather than red blood cell counts, into the apparatus of the present invention by means of the thumb wheel 30 shown in FIG.

Figure 8 shows the slight circuit changes to accomplish this. The transducer 10 and the AC amplifier 12 are the same as those shown in FIG.
Then, the output A′ of the AC amplifier 12, that is, BLRI is the second
The signal is input to the baseline return circuit 101 shown in Figure A.
The output of the recovery circuit is BLRO, as described above, which is connected to peak hold circuit 131 shown in FIG. 2A to generate signal E _p .

As shown in FIG. 5, E _p is a signal whose magnitude represents a platelet peak or a red blood cell peak. According to this modification of the invention, rather than coupling E _p to the circuit of FIG. 2B, a hematocrit pulse converter unit 201 is provided which receives the E _p waveform and connects it to its output line 33'. This pulse train increments the pulse counter 28 (FIG. 1). Of course, this pulse counter counts hematocrit pulses, not red blood cell pulses. Thumb-operated wheel switch 30' contains known hematocrit data of the blood sample under test, and according to the invention, when pulse counter 28 reaches the value of thumb-operated wheel switch 30', a stop signal is generated. Stop the platelet counter 24. Digital reader 26 then provides a platelet count per unit volume.

So, generally speaking, the only major change in the existing circuit is to effectively integrate the red blood cell pulse represented by E _p to give the required percent volume units for known standard hematocrit data. In this case, a hematocrit pulse converter 201 is used. To give proper timing, indicate the presence of red blood cells rather than platelets (E
_p represents either) signal ST from unit 121 of FIG. 2A, along with a signal such as that available from line B of FIG. -1 is used by this pulse converter.

Therefore, with a simple set of switches (not shown), the device of the present invention can convert red blood cell counts (particles per unit volume) to hematocrit (percentage). Such a switch is conveniently connected to a thumb-operated wheel 30.

FIG. 9 shows unit 201 of FIG. 8 in more detail. The signal E _p is arithmetic amplifier 202
The amplifier is enabled at its negative inverting input by the output from NAND gate 203. The output from the NAND gate 203 is generated only when the timing pulse ST1 (FIG. 5) and the red blood cell presence signal match. The width of the selected E _p portion is of course determined by ST1 and is, for example, about 5 microseconds.

The output of amplifier 202 drives a variable current source consisting of transistor 203 and resistor R. The collector of this transistor is connected to a storage capacitor C. The charge or current generated by this current source is E _p divided by R and is designated I _A . The level of charge on capacitor C is monitored by a comparator 204 having a reference input. When this level exceeds the reference input, the D-type flip-flop 205 is activated, and via the agate 206
Output is generated to. The other input of NOR gate 206 is controlled by the output of NAND gate 203, so NAND gate 203
Controls the output of 6. Normally, this output clamps the constant current source _ID via diode 207. However, when the current source is unclamped during the ST1 timing period, this current source ID acts as a charge imparting means, discharging the capacitor C towards the level corresponding to the reference of the comparator 204. Let's put it back.

The circuit of FIG. 9 therefore acts as an integrator for the peak of the red blood cell pulse indicative of hematocrit. More importantly from an operational point of view, the common timing means is the analog input voltage means 202.
and the charging current _ID , such that they are enabled only during a common time interval of fixed width. This considerably improves the accuracy of the circuit.

From an operational point of view, the amount of charge stored in capacitor C is determined by the time T of signal ST1 and I _A (i.e. E
_p /R). This is I _D ×T
balanced by a digital charge pulse feedback circuit that generates _NH , where _NH is the number of hematocrit pulses on line 33'. The peak of the red blood cell pulse is then integrated (percentage) and converted to digital form.

Additionally, by adjusting the circuit values, the ratio of actual red blood cell pulses compared to one hematocrit pulse generated on line 33' may be 10:1.
It is. In other words, several red blood cell pulses are required to charge capacitor C to a charge level that exceeds the reference value of comparator 204 and generates a hematocrit pulse. Therefore, the maximum peak value of E _p will probably give only one hematocrit pulse, for example. Such an arrangement considerably reduces errors in this type of conversion system.

[Brief explanation of the drawing]

1 is a schematic diagram showing a particle ratio calculation device suitable for carrying out the invention for measuring platelet density in blood; FIG. 2 is a diagram of FIGS. 2A, 2B, 3 and 3; Figure 4 shows how to combine the 2nd
Figures A and 2B are schematic diagrams showing a preferred embodiment of the pulse generation portion of the present invention; Figure 3 is a schematic diagram of the counter and display portion of the device shown in Figure 1; and Figure 4 is a schematic diagram of the red blood cell counter of the present invention. FIG. 5 is a timing diagram showing the timing relationships of various waveforms corresponding to FIGS. 2A and 2B; FIGS. FIG. 8 is a block diagram of another embodiment of the invention, and FIG. 9 is a detailed view of FIG. 8. 10... particle sensing transducer, 12...
AC amplifier, 14...Baseline return circuit, 16...High threshold detector, 18...Low threshold detector, 20...Peak detector, 22...Pulse classification unit, 24... Platelet pulse counter, 26... Platelet display unit, 28... Red blood cell pulse counter, 30... Thumb operated wheel switch array, 33, 35...
Pulse line, 101...Baseline return circuit, 111
...Low threshold detector, 121...
Strobe sequencer circuit, 131...Peak holding circuit, 161...Peak detector, 171...High threshold detector, 181...Pulse classification unit.

Claims (1)

[Claims] 1 The concentration of particles of the first type suspended in a fluid that suspends at a predetermined concentration a second type of particles that differ in detectable physical properties from the first type of particles. A measuring device is disposed in a flow of said fluid or a homogeneous dilution of said fluid and senses said each type of particle to differ in electrical properties corresponding to said physical property of each type of particle. A particle sensor that generates electric pulses corresponding to each type of particle, a discrimination means that electrically discriminates the electric pulses corresponding to each type of particle from the particle sensor, and a discrimination means that generates electric pulses corresponding to each type of particle. a first counting means for counting the number of the electric pulses corresponding to the first type of particles discriminated and generating a first count value; a second counting means for counting the number of the electric pulses corresponding to the second type of particles in the fluid to generate a second count value; means for detecting the first count value when a value corresponding to a predetermined concentration is reached and generating an output representative of the concentration of particles of the first type from the detected first count value; A device characterized by comprising: